Lipid particles and suspensions and uses thereof

ABSTRACT

The present invention relates to formulations and methods for the mucosal and parenteral administration of lipid particles an suspensions. The formulations of this invention are stable lipid particles useful for oral delivery of water-insoluble therapeutic agents, vaccines and diagnostics. The compositions of this invention promote the mucosal absorption of biologically active molecules across mucosal epithelial barriers. Stabilization of lipid particles is achieved by coating the hydrophobic central core with a polymer shell. The polymer shell can include bioadhesive agents, ligands, and absorption promoting agents. This invention relates to oral drug delivery systems for hydrophobic drugs, and in particular is concerned with improving the bioavailability of hydrophobic drugs from such systems. Using this system, anticancer drugs such as taxanes are orally effective.

BACKGROUND

1. Field of the Invention

The present invention relates to delivery systems for the mucosal andparenteral administration of biologically active molecules, including,but not limited to, therapeutic agents, vaccines, allergens, antigensand diagnostic agents. In particular, the present invention relates tolipid suspension and lipid particle compositions comprising lipids,polymerized lipids and derivatives thereof, fatty acid esters andderivatives thereof, additional water miscible solvents and surfactants,and optionally one or more biologically active agents, and methods ofadministering biologically active molecules to an animal utilizing saidcompositions. The compositions of this invention further comprisepolymeric ingredients to promote stability in the gastrointestinal tractand in contact with mucosal fluids. The compositions of the inventionpromote the absorption of biologically active molecules across mucosalepithelial barriers and are especially suitable for promoting theabsorption of poorly water-soluble drugs, such as paclitaxel and othertaxanes. The compositions of the invention can be used therapeutically,diagnostically or cosmetically.

2. Background of the Invention

Many drugs are limited in their development by the parenteral route ofadministration. Thus one of the great challenges in the improvement ofthe therapeutic potential of poorly water-soluble drugs is thedevelopment of systems that will provide optimal solubility and oral ormucosal bioavailability. Drugs, compounds, biologically active agentsand the like with little or no solubility in water are also referred toas lipophilic or hydrophobic and these terms are indistinguishablewithin the scope of the present invention.

The most convenient way to administer drugs into the body is by oraladministration. However, many drugs, in particular water-insoluble drugsand macromolecular compounds such as proteins and peptides, are poorlyabsorbed or unstable during passage through the gastrointestinal (GI)tract. The administration of these poorly absorbed or unstable drugs isgenerally performed through parenteral injection. Many of the morerecently developed anticancer agents, though effective, can only beadministered via intravenous injection because of low solubility inwater. A large proportion of the macromolecular drugs developed byrecombinant DNA methods can be delivered only by injection of themolecules, either subcutaneously or through intravenous administration.

Lipid systems have been widely exploited for development of drugdelivery vehicles and systems. Most of the lipid-based particulatesystems that have been developed for delivery of poorly water-soluble,or lipophilic drugs, are micellar, emulsion or suspension typeformulations. For many hydrophobic drugs, there remains the need to finda carrier system that will enhance the bioavailability of such drugs inthe GI tract.

One approach to making suitable formulations of water-insoluble drugs isto solubilize a hydrophobic therapeutic agent in an oil and dispersethis oil phase in an aqueous phase. Depending on whether an oil is asolid or liquid at the ambient temperature, the oil-in-water emulsioncan be characterized as a solid lipid particulate. The dispersion may bestabilized by emulsifying agents and provided in emulsion form. In awater milieu, drugs dissolved in the oil phase or the solid lipid corecan be dispersed by mechanical force to create microdroplets ormicrospheres that are stable in storage as a pharmaceutical preparation.The formation of a stable oil-in-water emulsion or lipid spheres dependson the use of surfactants that form the interface between the strictlyhydrophobic oil and water. Depending on the nature of the oil andco-surfactant, either large droplets or particles are formed. Furthercontrol over size of droplets or particles can be obtained by highpressure homogenization or similar shear forces. Lipid particles aretypically formed at higher ambient temperatures to melt the hydrophobiccomponents.

Oil-in-water (o/w) emulsions are also commonly formed from oil(s),surfactant(s), and an aqueous phase. Typically, oils are used in drugdelivery systems to solubilize lipophilic drugs and to make them moreeffective and less toxic. Oils used in typical emulsions are any of anumber of oils such as mineral, vegetable, animal, essential andsynthetic oils, or mixtures thereof. In many cases oils rich intriglycerides, such as safflower oil, cottonseed oil, olive oil orsoybean oil are used. Such emulsions contain the hydrophobic therapeuticagent solubilized in an oil phase that is dispersed in an aqueousenvironment with the aid of a surfactant or a combination ofsurfactants. Therefore, one approach to making suitable formulations ofhydrophobic drugs is to solubilize a hydrophobic drug in an oil and todisperse this oil phase in an aqueous phase.

For many hydrophobic therapeutic agents, solubility is too low inaqueous solution to offer formulations that can deliver therapeuticallyeffective doses. Hydrophobic therapeutic agents, while poorly soluble inaqueous solution, could be sufficiently lipophilic that therapeuticallyeffective concentrations can be prepared in triglyceride-based solvents.The colloidal oil particles sizes are relatively large, ranging fromseveral hundred nanometers to several microns in diameter, in a broadparticle size distribution. Although triglyceride-based pharmaceuticalcompositions are useful in solubilizing and delivering some hydrophobictherapeutic agents, such compositions are subject to a number ofsignificant limitations and disadvantages. Under most conditions,emulsions are thermodynamically unstable, the droplet spontaneouslyagglomerating, eventually leading to complete phase separation. Thetendency to agglomerate and phase separate presents problems of storageand handling, and increases the likelihood that pharmaceutical emulsionsinitially properly prepared will be in a less optimal, less effective,and poorly-characterized state upon ultimate administration to apatient.

In other cases, therapeutic compounds, although hydrophobic, areinsufficiently soluble in tri-glycerides and cannot be formulated solelyin triglyceride oils. Surfactants are also required to form solid lipidsuspension. And the same forces that operate in liquid oil phase alsocause the precipitation of hydrophobic drugs at the interface of lipidswith water upon short or long term storage and destabilize lipidparticle suspension systems.

Although many lipid-based systems are used to promote absorption ofdrugs with poor water solubility, lipid-based systems still have severaldrawbacks in the development of orally or mucosally delivered drugs.Most lipid systems comprise triglycerides which are readily digested inthe milieu of the digestive system. Upon digestion of the triglycerides,the encapsulated hydrophobic therapeutic agent may precipitate and maynot be absorbed. If absorption of the hydrophobic therapeutic agentoccurs, the digestion of the triglycerides also causes rapid rather thansustained absorption.

Lipid-based delivery systems such as emulsion and microemulsionssystems, or lipid particulate systems are based on the use of polarlipids and related amphiphilic surfactant molecules to control theinteraction of hydrophobic molecules with water. In many cases, deliverysystems for hydrophobic drugs have also required the inclusion oforganic solvents that are water miscible in order to increase themolecular interactions between drugs and lipid or surfactant components.

Lipid-based delivery systems may additionally incorporate absorptionenhancers, such as the salicylates, bile salts and other surfactants.Absorption enhancers may function to increase the permeation of peptide,protein, and lipophilic molecules across epithelial barriers because oftheir interaction with the GI mucosa and concomitant opening of thetight junctions. A wide variety of amphiphilic molecules are known tobehave as absorption enhancers. In addition, bile salts and salicylates,medium chain fatty acid salts and esters, and medium chainmonoglycerides and di-glycerides are known to have mucosal absorptionenhancing activity. Absorption enhancement with these molecules isattributed to the presence of medium chain C₆-C₁₂ fatty acyl chains(6-12 carbon atoms in length), particularly those esterified with C₈-C₁₀fatty acids (8-10 carbon atoms in length).

Lipids and surfactants are differentiable from short and long chainhydrocarbons in that they are amphiphilic molecules, having bothhydrophilic and hydrophobic moieties. Surfactants are convenientlyclassified on an empirical scale known as the hydrophile-lipophilebalance (HLB) which runs from about 1 to about 45 for ionic surfactantsand from about 1 to about 20 for non-ionic surfactants. HLB valuescloser to 1 represent surfactants with more lipophilic character, whileHLB values that are greater than about 10 represent more hydrophilicsurfactants.

Most familiar in the class of lipid vehicles are liposomes. Liposomesare traditionally formed from pure or mixed phospholipids or mixtures ofphospholipds with cholesterol or fatty acids. The characteristic featureof liposomes is the formation of an interfacial bilayer membrane thatseparates an internal water compartment from the external water milieu.Drugs and other active materials can be entrapped within the internalaqueous space. Conventional liposomes have been used successfully todevelop commercial pharmaceutical compositions that abrogate thetoxicity of certain drugs such as amphotericin, when administeredintravenously. A major problem encountered with the development ofliposomes as drug delivery vehicles is their poor ability to withstandexposure to stomach acids, bile salts and phospholipases. Fordevelopment of stable lipid drug delivery vehicles, polymerized liposomesystems have been developed. Most of the work with polymerized lipidparticles has been focused on polymerized liposomes, where apolymerizable phospholipid or fatty acid is incorporated in the lipidbilayer of the liposome which upon polymerization produced a crosslinkedbilayer with increased rigidity and physical stability.

U.S. Pat. No. 5,160,740 (Hasegawa, E., et al.) discloses polymerizationof a 3,5 polymerizable 2,4-diene phospholipid, cholesterol, and apolymerizable 2,4-diene fatty acid to form a polymerized macromolecularendoplasmic reticulum. Additionally, U.S. Pat. No. 5,762,904 (Okada, J.,et al.) discloses the use of polymerized liposomes for the delivery oforal vaccines. Polymerized liposomes are formed using a bilayer formingphospholipid or mixtures of said phospholipids with non-phospholipidstructures. The presence of the polymer phospholipid results in amembrane that resists dissolution by detergents and bile salts and ismore acid resistant. A number of additional polymerizable phospholipidsare described in Regan, in Liposomes: from Biophysics to Therapeutics(Ostro, ed., 1987), Marcel Dekker, N.Y.

Incorporating a targeting ligand on the surface of a liposome mayincrease the efficiency of absorption of drugs encapsulated in thoseliposomes. U.S. Pat. No. 6,004,534 (Langer, R. S. and Chen, H.)discloses modifications to the surface of polymerized liposomes in whichplant lectins were conjugated. Such lectins recognize receptors on thesurface of epithelial cells and promote greater adherence of theliposomes to M cells (Chen et al., 1996, Pharmaceutical Research13:1378-1383).

Additional strategies to enhance the bioavailability of hydrophobicdrugs include methods to increase surface area of drug crystals and theco-inclusion of P-glycoprotein (PGP) inhibitors in formulations in aneffort to increase absorption. Many drugs are substrates for the PGP,which acts as an efflux pump. As disclosed in U.S. Pat. No. 6,245,8,05(Broder, S., K. L. Duchin, and S. Selim/Baker Norton Pharmaceuticals,Inc.), cyclosporin A may be used to enhance the bioavailability ofhydrophobic drugs by inhibiting PGP. Additional compounds that are knowninhibitors of PGP can also be used to enhance the bioavailability oflipophilic drugs. These include other PGP inhibitors such ascyclosporine analogues, surfactants such as poloxamers, polysorbates,α-tocopherol polyethylene glycol esters, as well as therapeutic agentsknown to affect the activity of PGP such as verapamil and ketoconazole.

U.S. Pat. No. 6,207,178 (K. Westesen) discloses solid lipid particlesand particles of bioactive agents and methods for the manufacture anduse thereof. A suspension formulation is manufactured by an emulsifyingprocess resulting in non-spherical particles (anisometric). These solidlipid particles provide a controlled release dosage form for poorlywater soluble drugs primarily by IV but also by peroral, nasal,pulmonary, rectal, dermal, and buccal route of administration. Waterinsoluble drugs can be melted in the lipid phase prior tohomogenization. The melted lipid phase is emulsified in the aqueousphase using high pressure homogenization or sonication. These particlescan be freeze dried by removing the liquid using ultrafiltration.

WO 00/06120 (Jeong, S., et al.) discloses lipid emulsion a and solidlipid nanoparticle as a gene or drug carrier. The emulsion is anoil-in-water lipid emulsion which is composed of non-triglyceride oilsand also solid lipid nanoparticles (SLN) composed of triglyceride orethyl stearate, phospholipids and non-ionic surfactants used as genetransfection agents and drug delivery systems. The SLN are preparedwithout melting of the lipids, by mixing the aqueous and the fattyphases.

U.S. Pat. No. 5,904,932 (T. DeVringer) discloses a preparationcontaining a suspension of solid lipid particles made of lipid andemulsifier for topical application.

U.S. Pat. No. 5,726,164 (Weder, H. G. and P. van Hoogevest) disclosesnanosuspensions for intravenous administration. Pharmaceuticalcomposition containing the active which is insoluble (specificallyN-benzoyl-staurosprin), a polyoxyethylene/polyoxypropylene blockcopolymer, purified lecithin, water soluble excipients glycerol andsorbitol along with ethanol and water as solvents for the IVadministration of insoluble drugs.

U.S. Pat. No. 5,188,837 (A. J. Domb) discloses lipospheres forcontrolled delivery of substances. A microsuspension containinglipospheres which are solid, water insoluble particles that have a layerof a phospholipid embedded on their surface. The lipid core is a solidsubstance to be delivered that is dispersed in wax.

For conventional oral delivery systems, the drug is released into the GItract within a short period of time, and plasma drug levels peak usuallywithin a few hours after dosing. A controlled release oral dosage formis designed to maintain drug levels at constant effectiveconcentrations. Generally, controlled delivery of lipophilic drugsrequires techniques different than those employed with hydrophilicdrugs. Lipophilic drugs must be solubilized in order to be released in acontrolled fashion.

Citation or identification of any reference in Section 2, or any sectionof this application shall not be construed as an admission that suchreference is available as prior art to the present invention.

SUMMARY OF THE INVENTION

The present invention is directed to a particle composition thatcomprises a hydrophobic phase, a surfactant, and a polymeric stabilizer.The composition may optionally comprise a biologically active agent. Thecomposition may also optionally comprise a solvent. The hydrophobicphase of the composition may comprise an oil or a mixture of oils. Apreferred oil is a triglyceride. Preferably the hydrophobic phasecomprises a triglyceride that is a liquid between 10° C. and 45° C., andmore preferably is a hydrophobic phase comprising a triglyceride that isliquid between 10° C. and 70° C.

The surfactant of composition may be alkyl glycerolphosphoryl choline, apolyoxyethylene polymer, a block copolymer of polyoxyethylene andpolyoxypropylene, or an ethoxylated glycerol ester. The surfactant mayalso comprise one or more glycerol fatty acid esters or hydrophilicderivatives thereof. Preferably the fatty acid esters have a length ofabout 6 to about 12 carbon atoms. In another embodiment, the surfactantmay comprise a monoglyceride, diglyceride, or a hydrophilic derivativeor analog thereof.

A preferred monoglyceride useful in the practice of the presentinvention, is a monoglyceride derivatized by a polyoxyethylene polymerof about 200 to about 10,000 daltons in molecular weight. Preferably thepolyoxyethylene polymer is from about 200 to about 4,000 in molecularweight. A preferred diglyceride useful in the practice of the presentinvention is a diglyceride derivatized by a polyoxyethylene polymer.Fatty acid esters useful in the present invention include fatty acidesters derivatized with acetic acid, citric acid, lactic acid, succinicacid, tartaric acid, or mixtures thereof. A preferred fatty acid esteruseful in the practice of the present invention is a caprylic acid orcapric acid. Preferred surfactants have a HLB value of about 1 to about45 and preferably from about 1 to about 20.

Solvents useful in the practice of the present invention include water,glycerol, sorbitol, mannitol, propylene glycol, ethylene glycol,polyethylene glycol, or mixtures thereof. In a preferred embodiment, thesolvent of the present invention comprises monoterpene or a derivativethereof. Preferred monoterpenes includes perillyl alcohol,perilladehyde, perillic acid, perillilic acid methyl ester andd-limonene.

The particles of the present invention may be suspended in an aqueousphase comprising an additive. Preferred additives include a suspendingagent, buffering agent, tonicity agent, an oxidizing agent, a reducingagent, an antimicrobial agent, a preservative, a stabilizing agent ormixtures thereof. In the compositions of the present invention, thehydrophobic phase is preferably present in an amount from about 0% toabout 50% by weight and more preferably in amount from about 10% toabout 25% by weight. Preferred biologically active molecules useful inthe compositions of the present invention include prophylactics ortherapeutic agents. They may also be a diagnostic agent. A preferredtherapeutic agent is a taxane or an analog thereof. A more preferredbiologically active molecule is paclitaxel.

In another embodiment of the present invention the biologically activemolecule is a topoisomerase inhibitor selected from the group consistingof etoposide, camptothecin, topotecan, or a derivative thereof. Incompositions of the present invention, the biologically active moleculeis preferably soluble at least 0.1 mg/ml in the hydrophobic phase, andmore preferably soluble at least 1.0 mg/ml in the hydrophobic phase. Thepolymeric stabilizers useful in the compositions of the inventioninclude but are not limited to natural polymers, synthetic polymers, ormixtures thereof. The polymeric stabilizers include polymerizable fattyacids or polymerizable phospholipids. The more preferred polymericstabilizers are ODP or OPS. Preferably the polymer may be formed byinterfacial ionic polymerization with water or other initiators.However, the polymer may also be formed by condensation ofcyanoacrylates, including but not limited to alkyl cyanoaciylates. Apreferred alkyl cyanoacrylate is ethyl 2-cyanoacrylate. The syntheticpolymer may also be a polylactide, a polyglycolide, a mixture ofpolylactide and polyglycolide, a polycaprolactone, a polyortho ester,polysebacic acid, polyfumaric acid, polyamides, polycarbonates,polyalkylenes, polyacrylamides, poly(hydroxy acids), polyanhydrides,polyortho esters, polyacrylate, polyvinyl alcohols, blends or copolymersthereof.

The particle composition of the pi-sent invention may also be in theform of a suspension. Preferably the size of the particles in thesuspension range from about 10 nm to about 10,000 nm and are preferablym the range of 10 nm to 1,000 nm. The present invention is also directedto methods for delivering biologically active methods to an animal,methods for treating, preventing, or ameliorating one or more symptomsof a disease in an animal, and methods for diagnosing a disease ordisorder in an animal using any of the foregoing compositions. Thecompositions of the present invention may be administered in the form ofa capsule, soft elastic gelatin capsule, caplet, aerosol, spray,solution, suspension, emulsion, sachet, tablet, powder, or granules. Thecomposition is preferably administered orally.

DETAILED DESCRIPTION OF THE INVENTION

Many systems for oral delivery of hydrophobic drugs are oil-basedwherein the hydrophobic drug being is dissolved in an oil. However, theadministration of a drug in oil alone is not advantageous because of thepoor miscibility of the oil with the aqueous environment of thegastrointestinal tract The present invention provides a stable lipidsuspension. The lipid suspensions of the present invention may be givento an animal alone or in combination with water-insoluble molecules,e.g. lipophilic drugs, for treating, preventing or diagnosing diseasestates. The lipid suspension of this invention is effective in promotingthe absorption of hydrophobic biologically active materials by mucosaltissues by protectively encapsulating one or more hydrophobic materialswithin a stable lipid particle comprised of one or more hydrophobicsolvents, one or more surfactants, and one or more polymericstabilizers. More than one drug or pharmaceutical agent and/orformulation at a time can be used according to the present invention toyield a desired pharmaceutical composition. Also, in accordance with thepresent invention is a method of treating disease, such as cancer, usinga drug delivery system for increasing the bioavailability of one or morehydrophobic drugs.

The lipid particles of this invention are stabilized by a polymer oroligomer shell, which encases the hydrophobic core of the liquidparticle. The polymer or oligomer shell is comprised of one or morepolymeric stabilizers which have undergone polymerization orcross-linking. The porosity of the polymeric shell is controlled by thedegree of polymerization or cross-linking of the polymeric stabilizers.The lipid particles and suspensions of the present invention provideimproved bioavailability of hydrophobic compounds because hydrophobicdrugs are contained in the central hydrophobic core of the lipidparticle and are prevented from contacting aqueous solution, such asgastrointestinal or mucosal fluid. In addition, the encapsulatedhydrophobic drugs are able to interact directly with the membranebarriers of absorptive cells. Bioavailability of hydophobic drugs canalso be increased by conjugating absorption promoting agents to thepolymeric compounds that form the shell of the lipid core. In additionto increase bioavailabilty, the polymeric shell provides increasedstability of lipid suspensions in aqueous storage milieu and increasedstability upon lyophilization. Absorption promoting compounds may becompounds that control the opening and closing of intercellular tightjunctions or that interact with epithelial cell receptors. In addition,absorption promoting compounds may be compounds that are bioadhesive ormucoadhesive.

Stabilization of the lipid particles can be achieved by forminghydrophobic biologically active molecule-containing particles in thepresence of monomeric polymerizable compounds and subsequentpolymerization in situ. Polymerization in situ results in a polymernetwork surrounding the central drug-containing core of the lipidparticle. Alternatively, stabilization of the hydrophobic core of alipid particle can be achieved by the addition of hydrophobic polymersthat interact with the hydrophobic moieties of the core-formingmaterials. The polymeric materials add physical rigidity to the systemby interacting with acyl side chains of the hydrophobic phase materialsto impede the dissolution of the hydrophobic phase by enzymes ordetergents in mucosal fluids. When polymerized, lipid particles havegreater stability in vitro upon contact with water or simulated oractual gastrointestinal fluid.

U.S. Pat. No. 6,187,335 (Brey, R. N., L. Liang) discloses polymerizablefatty acid compounds. These compounds are aliphatic fatty acids withpolymerizable groups in the head group or in the aliphatic chain. Suchfatty acids are further modifiable by extension of their hydrophilichead groups by ethylene glycol addition or addition of other hydrophilicgroups. The structure of these fatty acids gives them uniquefunctionality and particular utility when used in conjunction with lipidparticles.

The fatty acids described by Brey and Liang are fully compatible withamphiphilic and lipid materials and when contacted with water formthermodynamically stable structures. When used with lipid particlesformed with oils as the principal hydrophobic phase, such fatty acidsform the outer shell with the polar head groups of the fatty acidoriented towards the water phase and the hydrophobic side chainscontained in the hydrophobic compartment. Polymerization of the fattyacids results in crosslinked lipid in which the fatty acid polymerstabilizes the lipid particle.

The surfactant group of the fatty acids is disposed between thepolymerizable group and the functional acid group, which can beoptionally omitted. The length of the polymeric chain of the surfactantgroup can be chosen to be short, medium or long, in order to control therelative hydrophilicity/hydrophobicity of the chain. A long-chainsurfactant group with significant hydrophilicity, for example, mayprovide hydrophilic groups that interact effectively with cellmembranes, may provide in and of itself the surfactant activity, or maycontain ligands that specifically bind with cellular receptors.

For the purpose of forming polymers in situ in the lipid particle orsuspension, polymerization of the fatty acid polymerizable moiety may becarried out using three methods: (1) actions of chemical initiators,e.g., redox pairs; (2) physical excitation, including sensitizedphotoinitiation, e.g., broad band UV or UV 360 nm or UV 302 nmirradiation, gamma-ray irradiation, cyanine dye with an argon laser; and(3) combination of chemical initiators with physical excitation. Methodsof initiating polymerization may result in harsh environment changes,such as large pH drops and inactivation of protein drugs. As a result,the most desirable method of polymerization initiation is wherepolymerization can be controlled and the activity of therapeuticmaterials is totally retained.

Precise control of polymerization level is sometimes difficult toachieve with the use of chemical initiators. In addition, additionalsteps are normally required to separate any unreacted initiators,especially when low level of polymerization is needed. Dienepolymerizable functions may be polymerized by exposure to short-wave ormid-wave ultraviolet light. Ultraviolet light at 302 nm may be used topolymerize diene function and damage to proteins and peptides can beminimized. Phenylacetophenone initiators combined with UV 360 nmirradiation has been used extensively in the polymerization of alkenofunctionalities, such as acrylated PEG hydrogel for biomedical andmolecular imprinting applications, polymethacrylate polymers forbiomaterials and tissue engineering, and styrene/acrylate/methacrylatenanoparticles for drug delivery. Long range UV wavelengths are usuallyoutside of the absorption range of proteins and would not cause damageto proteins.

One embodiment of this invention is a lipid particle comprising one ormore hydrophobic solvents, one or more surfactants, one or morebiologically active hydrophobic molecules, and a polymeric shell, saidpolymeric shell comprised of one or more polymeric stabilizers. Apreferred hydrophobic solvent is an oil. The lipid particle of thepresent invention is a spherical or non-spherical particle with adiameter preferably between about 1 nm to about 1000 nm, and morepreferable with a diameter between 10 nm and 1000 nm, and mostpreferably with a diameter between 50 nm and 500 nm.

The stable lipid particle according to the present invention comprisesone or more hydrophobic solvents, preferably an oil that is a liquid atroom temperature or an oil that is a liquid at body temperature orabove. A drug or pharmaceutical with poor solubility in water may besolubilized in the hydrophobic solvent and can subsequently be dispersedin an aqueous phase for administration to an animal.

In a preferred embodiment of the invention, the hydrophobic solvent ispresent in an amount of about 5% to about 50% of the total weight of thelipid particle. In a more preferred embodiment of the invention, thehydrophobic solvent is present in an amount of about 10% to about 40% ofthe total weight of the lipid particle. In the most preferred embodimentof the invention, the hydrophobic solvent is present in an amount ofabout 20% to about 30% of the total weight of the lipid particle.

The lipid particle according to the present invention comprises one ormore surfactants. The surfactants of the current invention can includeemulsifying agents with HLB values from 1-45, from 1-20, from 5-20, from5-15. Typically, surfactants or emulsifiers within HLB in the range of5-20 can be used. In the present invention, the preferred HLB range forthe surfactant(s) is between approximately 5 and 15. Preferredsurfactants utilized in accordance with the present invention includelecithn. Other preferred surfactants include ethoxylated derivatives ofmedium chain mono- and di-glycerides such as Labrasol (caprylic/capric(C₈/C₁₀) polyethylene glycol mono- and di-glycerides, GattefosseCorporation). Any suitable surfactant may be employed alone or incombination with other surfactants. For example, egg yolk phospholipidssuch as lecithin or polyethylene oxide-polypropylene oxide blockcopolymers (Pluronics) such as Pluronic F68 (Poloxamer 188), having amolecular weight of about 8,000. Other suitable Pluronics, includePluronic F87 (Poloxamer 237) with an average molecular weight of about7,500 and Pluronic F127(Poloxamer 407) with an average molecular weightof about 12,000. Ethoxylated diacyl glycerol and dialkyl ether glycerolare useful surfactants. The lipid particles of this invention maycontain alkylphosphoryl choline or alkylglycerophosphoryl choline andother lipid surfactants such as 1,2-dioctylglycero-3-phosphoryl choline,1,2-ditetradecylglycero-3-phosphoryl choline,1,2-dihexadecylglycero-3-phosphoryl choline,1,2-dioctadecylglycero-3-phosphoryl choline,1-hexadecyl-2tetradecylglycero-3-phosphoryl choline,1-octadecyl-2-tetradecylglycero-3-phosphoryl choline,1-tetradecyl-2-octadecylglycero-3-phosphoryl choline,1-hexadecyl-2-octadecylglycero-3-phosphoryl choline,1-2-dioctadecylglycero-3-phosphoryl choline,1-octadecyl-2-hexadecylglycero-3-phosphoryl choline,1-tetradecyl-2-hexadecylglycero-3-phosphoryl choline,2,2-ditetradecyl-l-phosphoryl choline ethane, and ethoxylatedderivatives of medium chain mono- and di-glycerides. Anionic surfactantswith HLB values preferably greater than about 10, such as alkyl or arylsulfates, sulfonates, carboxylates or phosphates, and cationicsurfactants with HLB values preferably greater than about 10, such asmono-, di-, tri- and tetraalkyl or aryl ammonium salts may also be usedin the practice of the present invention. Non-ionic surfactants such aspolysorbate 80 (Tween 80) and polyalcohols such as polyvinyl alcohol mayalso be used. Zwitter-ionic surfactants that have a combination of theanionic or cationic groups, and whose hydrophobic part consists of anyother polymer, such as polyisobutylene or polypropylene oxides, may alsobe used. Mixtures of these surfactants may also be used as may othersurfactants well known in the art. The surfactant of this invention hasHLB values preferably from 1-20, more preferably from 5-20, and evenmore preferably from 5-10 or from 10-15.

In a preferred embodiment of the invention, the surfactant is present inan amount of about 15% to about 90% of the total weight of the lipidparticle. In a more preferred embodiment of the invention, thesurfactant is present in an amount of about 10% to about 30% of thetotal weight of the lipid particle. In the most preferred embodiment ofthe invention, the surfactant is present in an amount of about 20% toabout 30% of the total weight of the lipid particle.

The lipid particle according to the present invention also comprises abiologically active hydrophobic molecule. Such hydrophobic moleculesinclude but are not limited to methotrexate, cis-platin and derivatives,vincristine, vinblastine, quinolone, ciprofloxacin, progesterone,teniposide, estradiol, doxorubicin, epirubicin, taxanes andtopoisomerase inhibitors. Other hydrophobic molecules useful in thepractice of the present invention include prostaglandins, amphotericinB, testosterone, beclomethasone and esters, vitamin E, cortisone,dexamethasone and esters, betamethasone valerete and other steroids,nifedipine, griseofulvin, cyclosporin, digoxin, itraconozole,carbamazepine, piroxicam, fluconazole, indomethacin, steroids,ibuprofen, diazepam, finasteride and diflunisal. Preferred topoisomeraseinhibitors include etoposide, camptothecin, topotecan, or derivativesthereof. Combinations of more than one hydrophobic drug orpharmaceutical ingredient with poor solubility in water may beformulated according to the present invention to yield a desiredpharmaceutical composition.

The lipid particle according to the present invention also comprises apolymeric shell. Such polymeric shell is a microstructure comprising oneor more polymeric stabilizers which have undergone polymerization and/orcross-linking. The polymeric shell is formed by means including, but notlimited to chemical initiation with oxidation reduction initiators, orhigh energy radiation such as gamma or ultraviolet irradiation, andcombinations thereof In a preferred embodiment of this invention, thepolymeric shell is formed by irradiation by ultraviolet light at either302 nm or 350 nm in the presence of chemical initiators.

Polymeric stabilizers preferably form microstructures such asmicroparticles, microtubules, microspheres, matrices, and microcrystalsthat are compatible with the hydrophobic phase of the surfactant mixturemay be used to stabilize the lipid particles and suspensioncompositions. Such nicrostructures encapsulate the lipid particle withintheir structure. A polymeric stabilizer may be a natural polymer, asynthetic polymer or a mixture thereof.

Polymeric stabilizers according to the invention include but are notlimited to polylactide, poly-glycolide, a mixture of polylactide andpolyglycolide, a hydrocarbon oligomer, a hydrocarbon polymer, apolycaprolactone, a polyortho ester, polysebacic acid, polyfumaric acid,a polyamide, a polycarbonate, a polyalkylene, a polyacrylamide,poly(hydroxy acid), a polyanhydride, a polyorthoester, blends andcopolymers thereof. Other polymeric stabilizers useful in the practiceof the invention include fatty acid or fatty acid derivatives orphospholipid or phospholipid derivatives that are polymerizable, 2,4octadecadienoic acid [ODA],2,4 octadecadienoyl-polyethylene glycol(200-4000) [ODP],2,4 octadecadienoyl-PEG (2004,000)-succinic acid [OPS],Bis-(2,4-octadecadienoyl)-polyethylene glycol (200-10,000) [BODP] andanalogs thereof. Appropriate analogs include analogs modified by singleamino acids or polypeptide chains, imido groups, polyamines, polyimines,polysaccharides, polyacids, polymers or co-polymer of propylene glycoland ethylene glycol. Other polymerizable moieties may also be used,including, conjugated dienes of C6-C24, conjugated diynes of C6-C24,methacrylate modified or sulfhydryl-containing polar groups orhydrophobic tails of the fatty acids. The use of 2,4 conjugated dienesresults in polymers that are linked to adjacent acyl groups in the sidechains in the internal hydrophobic phase, where use of sulfhydrylcontaining polymerizable fatty acids results in head grouppolymerization at the interface of the aqueous and hydrophobic phases.

In addition, other polymerizable fatty acid derivatives can be used tostabilize the lipid particles. The polar head group, for example, mayconsist of amino acids, polypeptides, polysaccharides, polyols,polyacrylic acids, polyimines, choline, peptidoglycols, glycopeptides,or other hydrophilic polymers with multiple positive or negativecharges. Further, compounds that are polymerizable fatty acidderivatives of glycerol or glyceryl phosphatidyl derivatives compatiblewith the hydrophobic lipid core may be used.

In a preferred embodiment of this invention, the polymeric stabilizer isDODPC (2,4 dioctacedadienoyl phosphatidyl choline). In another preferredembodiment of this invention, a second polymeric stabilizer is apolymerizable fatty acid with polyethylene glyccol polar head groups ora polymerizable phopholipid. In a more preferred embodiment of thisinvention, the polymeric stabilizer ODP (2,4octadecadienoyl-polyethylene glycol −200-4000).

In another preferred embodiment of this invention, the polymeric shellcomprises a polymerizable fatty acid monomer or derivative, or monomerswhich have undergone interfacial ionic polymerization with water, suchcondensation of cyanoacrylates, alkylcyanoacrylates (e.g. ethyl2-cyanoacrylate).

In addition to the above constituents, the lipid particle of thisinvention may contain a secondary surfactant or “co-surfactant”. Suchsecondary surfactants include but are not limited to any of thesurfactants described above, as well as Labrasol (GattefosseCorporation), which is comprised of a mixture of capric caprylic(C₈-C₁₀) mono- and di-glycerides triglycerides. The secondary surfactantof this invention has HLB values preferably from 5-20, more preferablyfrom 5-15, and even more preferably from 5-10 or from 10-15.

In a preferred embodiment of the invention, the secondary surfactant ispresent in an amount of about 16% to about 89% of the total weight ofthe lipid particle. In the most preferred embodiment of the invention,the secondary surfactant is present in an amount of about 10% to about40% ofthe total weight of the lipid particle.

In addition to the above constituents, the lipid particle of thisinvention may contain a second hydrophobic solvent, which is miscible inthe hydrophobic solvent described above. The second hydrophobic solventmay further solubilize the hydrophobic biological agent Such secondhydrophobic solvents include but are not limited to polyethylene glycol,glycerol and related esters of fatty acids, polymerizable fatty acids,or polymerizable lipids, and monoterpene alcohols such as perillylalcohol or lirnonene.

In addition to the above constituents, the lipid particle of thisinvention may contain a hydrophobic polymer. Hydrophobic polymers areinsoluble in water and soluble in organic solvents. Such hydrophobicpolymers include, but are not limited to polymers comprising polylacticacid, polyglycolic acid, polyorthoesters, polysebacic acid, polymethylmethacrylate, polyacrylates, polystyrenes, and polyfumarate. Thealiphatic chains of hydrophobic polymers interact primarily with thehydrophobic side chain of oils and surfactants of the hydrophobic coreof the lipid phase to form a loose network of polymer chains therebystabilizing the hydrophobic core of a lipid particle.

In addition to the above constituents, the lipid particle of thisinvention may contain, monomeric compounds. Such monomeric compoundsinclude but are not limited to members of the cyano-acrylate family,such as 2-cyanoacrylate (ECA). Monomeric compounds that undergopolymerization in contact with water may be used to create polymers atthe interface of the aqueous phase and the hydrophobic phase. Forexample, a solution of 2-cyanoacrylate (ECA) dissolved in methylenechloride can be added to the hydrophobic phase of a preformed lipidcore. Upon stirring, ECA contacts the aqueous phase and polymerizationis initiated. The removal of solvent by evaporation results inpolymerization of ECA into a polymer principally at the interface of thelipid particle and water interface.

In addition to the above constituents, the lipid particle of thisinvention may contain other lipidic compounds. Such lipidic compoundsinclude, but are not limited to, low melting temperature waxes,including N.F. White Beeswax, Soy wax, Carnuba wax, Castor wax,Microwax, and other such waxes. Such waxes can be melted and mixeddirectly with polymeric stabilizers such as mono- or di-glyceride fattyacid esters.

The lipid particles and suspensions according to the present inventionalso comprise a hydrophobic therapeutic agent. Such hydrophobictherapeutic agents include but are not limited to methotrexate,cis-platin and derivatives, vincristine, vinblastine, quinolone,ciprofloxacin, progesterone, daunorubicin, teniposide, estradiol,doxorubicin, epirubicin, and taxanes. Other hydrophobic therapeuticagents useful in the practice of the present invention includeprostaglandins, amphotericin B, testosterone, beclomethasone and esters,vitamin E, cortisone, dexamethasone and esters, betamethasone valereteand other steroids, nifedipine, griseofulvin, cyclosporin, digoxin,itraconozole, carbamazepine, piroxicam, fluconazole, indomethacin,steroids, ibuprofen, diazepam, finasteride and diflunisal. Othertherapeutic agents may also be used including antibiotics (antiviral,antibacterial, antihelminthic, antiplasmodial, or antimycotic),analgesics and local anesthetics, antidepressants, antipsychotics,sedatives, hypnotics, hormones, cytokines, vaccine adjuvants andantigens, immunosuppressive agents, vasodilators, antiarrhythmics,calcium antagonists, cardiac glycosides, oligonucleotides,oligopeptides, anti-emetics, and migraine therapeutics.

The lipid particles and suspensions according to the present inventionmay also comprise hydrophilic therapeutic molecules that can bederivatized with a hydrophobic compound. Upon derivatization with ahydrophobic compound, the hydrophilic molecule may be solubilized withinthe hydrophobic phase of the lipid particle. Methods of derivatizationinclude but are not limited to conjugation of fatty acids through esterlinkages to the amino terminal amino acid of a peptide or to epsilonamino groups of lysines resulting in esterification of acyl chains toproteins and peptides. Further, oligosaccharides and polysaccharides maybe derivatized through available hydroxyl groups and both DNA and RNAmay be selectively acylated using similar techniques. The derivatizingagent may include a number of hydrophobic acyl groups.

Hydrophilic therapeutic and bioactive molecules may also be physicallyassociated with the surface of a lipid particle through ionicinteractions. An lipid particle with a net positive surface charge maybe made using amphipathic surfactants comprising positively chargedfatty acid chains. Such positively charged lipid particles will readilyadsorb nucleic acids and other negatively charged compounds to thesurface of the particle. Lipid particles with surface adsorbed DNA maybe used for gene transfection vehicles in vitro and gene transfer agentsfor treating genetic diseases and for genetic vaccination.

A more preferred embodiment of the present invention includes a lipidparticle comprising one or more hydrophobic solvents, one or moresurfactants, one or more biologically active hydrophobic molecules withanti-cancer activity, and a polymeric shell, said polymeric shellcomprised of one or more polymeric stabilizers. Lipid particles of thismore preferred embodiment are useful for the administration oflipophilic anti-cancer agents. Simultaneous delivery of combinations ofanti-cancer agents increases the benefit over monotherapies, and thusmore effectively treats tumors. Furthermore, the combination of multiplehydrophobic anti-cancer agents may lead to synergistic anti-canceractivities.

One of the preferred anti-cancer agents useful in the lipid particle ofthe present invention is etoposide. Another of the preferred anti-canceragents useful in the emulsions of the present invention is doxorubicinand its lipophillic derivatives thereof. Other preferred anti-canceragents include daunorubicin, irenotecan, mitomycin, bleomycin,procarbazine, altretamine, and lipophilic pro-drug derivatives ofmethotrexate, hydrophobic cis-platin derivatives such as2-hydrazino-4,5-dihydro-1H-imidazole with platinum chloride or5-hydrazino-3,4-dihydro-2H-pyrrole with platinum chloride, vincristine,vinblastine, teniposide, epirubicin, camptothecin, teniposide,topotecan, etoposide, teniposide, monophosphoryl Lipid A, and muramyldipeptide derivatives. Still other preferred anti-cancer agents usefulin the practice of the present invention are taxanes, including but notlimited to lipid-soluble taxane and taxane derivatives includingpaclitaxel; docetaxel; spicatin; taxane-2,13-dione, 5β-, 9β-,10β-trihydroxy-, cyclic 9,10-acetal; taxane-2,13-dione, 5β-, 9β-,10β-trihydroxy-,cyclic 9,10-acetal; taxane-2β-, 5β-, 9β-, 10β-tetrol,cyclic 9, 10-acetal; cephalomannine-7-xyloside;7-epi-10-deacetylcephalomannine; 10-deacetylcephalomannine;cephalomannine; taxol B; 13-(2′,3′-dihydroxy-3′-phenylpropionyl)baccatin III; yunnanxol; 7-(4-Azidobenzoyl)baccatin III;N-debenzoyltaxol A; O-acetylbaccatin IV; 7-(triethylsilyl)baccatin III;7,10-Di-O-[(2,2,2-trichloroethoxy)carbonyl]baccatin III; baccatin III13-O-acetate; baccatin diacetate; baccatin; baccatin VII; baccatin VI;baccatin IV; 7-epi-baccatin III; baccatin V; baccatin I; baccatin III;baccatin A; 10-deacetyl-7-epitaxol; epitaxol; 10-deacetyltaxol C;10-deacetyl-7-xylotaxol; 7-epi-10-deacetyltaxol; 10-deacetyltaxol;taxagafme and 10-deacetyltaxol B.

In a preferred embodiment of this invention, paclitaxel is present in anamount of about 0.1% to about 20% by weight of the lipid particle.

Another embodiment of the present invention is a lipid suspensioncomprising an aqueous solvent and a lipid particle, said lipid particlecomprising one or more hydrophobic solvents, one or more surfactants,one or more biologically active hydrophobic molecules, and a polymericshell, said polymeric shell comprised of one or more polymericstabilizers. A preferred hydrophobic solvent is an oil. The lipidsuspension of the present invention contain a stable discrete lipidparticle entity in a hydrophilic milieu such as water, where the lipidparticle protectively encapsulates one or more solubilized hydrophobicagents.

The aqueous solvent may further comprise drugs or pharmaceuticals whichare soluble in aqueous solution. Combinations of drugs orpharmaceuticals in the hydrophobic phase and the aqueous phase may beformulated to yield a desired pharmaceutical composition.

In addition to the above constituents, the lipid suspension of thisinvention may include other pharmaceutically acceptable compounds orexcipients to increase the stability of the lipid particles insuspension systems. The lipid suspension of this invention may include asuspending agent to disperse lipid particle evenly in an aqeuous milieuand to prevent the solid lipid particles from separating from theaqueous phase. Such pharmaceutically acceptable compounds or excipientsinclude but are not limited to Xanthan Gum, tragacanth, cetyl alcohol,stearic acid, and/or beeswax (Remington's Pharmaceutical Sciences,1975). Other preferred suspending agents include carboxymethylcellulose,methylcellulose, microcrystalline cellulose, poly(vinylpyrrolidone), andbentonite. In addition, tonicity and buffering agents to control pH ofthe suspending solution, as well as flavoring and coloring agents may beadded.

Another embodiment of the present invention is a vaccine lipidsuspension comprising an aqueous solvent and a lipid suspension, saidlipid suspension comprising one or more hydrophobic solvents, one ormore surfactants, one or more hydrophobic antigens, and a polymericshell, said polymeric shell comprised of one or more polymericstabilizers. A preferred hydrophobic solvent is an oil.

The vaccine lipid suspension of the present invention preferably furthercomprises one or more pharmaceutically acceptable excipients which arecompatible with the said antigen. Suitable excipients include but arenote limited to water, saline, dextrose, glycerol, ethanol, orcombinations thereof.

The vaccine lipid suspension of the present invention preferably furthercomprises auxiliary substances including but not limited to wetting oremulsifying agents, and pH buffering agents.

The antigen of the present invention may be formulated into the vaccinelipid suspension as a neutral or salt form. Pharmaceutically acceptablesalts include but are not limited to the acid addition salts (formedwith free amino groups), which are formed with inorganic acids,including but not limited to hydrochloric or phosphoric acids, ororganic acids such as acetic, oxalic, tartaric and maleic. Salts formedwith free carboxyl groups are preferably derived from inorganic bases,including but note limited to sodium, potassium, ammonium, calcium, orferric hydroxides, and such organic bases as isopropylamine,trimethylamine, 2-ethylamino ethanol, histidine, and procaine.

The vaccine lipid suspensions of the present invention may bemultivalent or univalent. Many methods may be used to introduce thevaccine formulations of the present invention; including but not limitedto oral, intradermal, intramuscular, intraperitoneal, intravenous,subcutaneous, intranasal, rectal, and via scarification (scratchingthrough the top layers of skin, e.g., using a bifurcated needle). Thepatient to which the vaccine is administered is preferably an animal,more preferably a mammal, most preferably a human, but can also be anon-human animal including but not limited to cows, horses, sheep, pigs,fowl (e.g., chickens), goats, cats, dogs, hamsters, mice and rats.

The vaccine lipid suspensions of the present invention comprise aneffective immunizing amount of one or more antigens and apharmaceutically acceptable carrier or excipient. Pharmaceuticallyacceptable carriers are well known in the art and include but are notlimited to saline, buffered saline, dextrose, Water, glycerol, sterileisotonic aqueous buffer, and combinations thereof. A further example ofphysiologically acceptable carrier is a physiologically balanced saltsolution containing one or more stabilizing agents including but notlimited to stabilized, hydrolyzed proteins and lactose. Thepharmaceutically acceptable carrier is preferably sterile.

The vaccine lipid suspensions of the present invention may be in theform of a liquid solution, suspension, emulsion, sustained releaseformulation, powder, and preferably solid forms such as capsules,tablets or pills. Vaccine lipid suspensions for oral administrationpreferably include standard carriers including but not limited topharmaceutical grades of mannitol, lactose, starch, magnesium stearate,sodium saccharine, cellulose, or magnesium carbonate.

Vaccine lipid suspensions in liquid form may be provided in ahermetically sealed container such as an ampoule or a sachet. Thevaccine formulations are generally stored at 4° C. prior to use. Theprecise dose of vaccine lipid suspension to be employed will depend onthe route of administration, and the nature of the patient, and shouldbe decided according to the judgment of the practitioner and eachpatient's circumstances according to standard clinical techniques.

An effective dose of the immunogenic antigen is that amount sufficientto produce an immune response to the antigen in the host to which thevaccine lipid suspension is administered. Use of purified antigens ascomponents of vaccine lipid suspensions may be carried out by standardmethods. If the immunogenic antigen is a protein, the purifiedprotein(s) should be adjusted to an appropriate concentration,formulated with any suitable vaccine adjuvant and encapsulated withinthe lipid suspension. Suitable adjuvants include but are not limited tomineral gels, such as aluminum hydroxide, surface active substances suchas lysolecithin or pluronic polyols, polyanions, peptides, oilemulsions, alum, Lipid A and derivatives of Lipid A, cytokines, and MDP.

Another embodiment of the present invention includes methods fortreating disease using a lipid suspension drug delivery systemcomprising administering to a mammal a lipid suspension in a suitableoral dosage form, such as a soft or hard-filled gelatin capsule, whereinsaid lipid suspension comprises an aqueous solvent and a lipid particle,said lipid particle comprising one or more hydrophobic solvents, one ormore surfactants, one or more biologically active hydrophobic molecules,and a polymeric shell, said polymeric shell comprised of one or morepolymeric stabilizers.

A more preferred embodiment of the present invention includes methodsfor treating cancer using a lipid suspension drug delivery systemcomprising administering to a mammal a lipid suspension in a suitableoral dosage form, such as a soft or hard gelatin capsule, wherein saidlipid suspension comprises an aqueous solvent and a lipid particle, saidlipid particle comprising one or more hydrophobic solvents, one or moresurfactants, one or more biologically active hydrophobic molecules withanti-cancer activity, and a polymeric shell, said polymeric shellcomprised of one or more polymeric stabilizers. The preferredanti-cancer agents are described above.

Another embodiment of the present invention includes methods fordiagnosing a disease using a lipid suspension drug delivery systemcomprising administering to a mammal a lipid suspension in a suitableoral dosage form, such as a soft or hard gelatin capsule, wherein saidlipid suspension comprises an aqueous solvent and a lipid particle, saidlipid particle comprising one or more hydrophobic solvents, one or moresurfactants, one or more biologically active hydrophobic molecules, anda polymeric shell, said polymeric shell comprised of one or morepolymeric stabilizers.

The lipid particles and lipid suspensions of the present invention arepreferably administered through mucosal tissue or epithelia The lipidparticles and lipid suspensions of the present invention are thereforeadministered by those routes which optimize uptake by mucosa, such assublingual, buccal, rectal and intranasal, and preferably oral. Thelipid particles and lipid suspensions of the present invention can bedelivered orally in the form of tablets, capsules, cachets, gelcaps,solutions, suspensions, topically in the form of creams, ointments,suppositories and the like, transdermally and parenterally.

If administered topically the lipid particles and lipid suspensions arepreferably administered in the form of an ointment or transdermal patch.If administered intranasally the lipid particles and lipid suspensionsare preferably administered in an aerosol form, spray, mist or in theform of drops. Suitable formulations can be found in Remington'sPharmaceutical Sciences, 16th and 18th Eds., Mack Publishing, Easton,Pa. (1980 and 1990), and Introduction to Pharmaceutical Dosage Forms,4th Edition, Lea & Febiger, Philadelphia (1985), each of which isincorporated herein by reference.

The lipid particles and lipid suspensions of the present invention aresuitable for administration to animals, preferably mammals and birds,and more preferably humans. For example, domestic animals such as dogsand cats, as well as domesticated herds, cattle, sheep, pigs and otherdomesticated mammals may be treated or vaccinated with the lipidparticles and lipid suspensions of the present invention. In a preferredembodiment, the lipid particles and lipid suspensions of the presentinvention are administered to humans lipid particles and lipidsuspensions are preferably provided in a hermetically sealed containersuch as an ampoule or sachet, and stored at 4° C. Dosages of the lipidparticle and suspension compositions will vary depending on theindividual patient and the mode of administration. Such dosages can bedetermined by a skilled physician using standard techniques.

The development of an oral formulation for an insoluble or poorlysoluble drug often involves the designing of a system that will affectthe pH of the micro-environment surrounding the drug form in the GItract after ingestion. In particular, the formulation may containdisintegrants and/or other agents that work to increase or decrease thepH of the micro-environment, and thus enhance drug dissolution. Inaddition, the drug may also be granulated to reduce its particle sizeand/or increase the surface area that is exposed to the gastric fluid.The amount of exposed surface area will affect the rate of drugdissolution and thus the amount of active drug that will be absorbed bythe patient. With respect to drug compounds of very poor or limitedsolubility, those skilled in the art have used co-solvents, surfactantsor wetting agents to reduce the surface tension of the liquidenvironment of the gastric fluid in which the active drug is to bedissolved. These agents wet the active drug more quickly so that more ofthe drug is exposed to the gastric fluid in a shorter time, and mayenhance its dissolution. Common types of surfactants and co-solventsthat can be used include the cationic, anionic (e.g., sodium laurylsulfate and gelatin), and nonionic types, as well as such co-solvents asthe polyethylene glycols (PEGs). The role of the binder in the tabletdrug form is to provide a tablet with sufficient hardness and integrity,but also must allow for sufficient disintegration and dissolution in thegastric environment. In this sense, a binder performs the oppositefunction of a disintegrant. The types of binders that can be used indrug formulations include gelatins of numerous grades, starches andstarch derivatives (including corn starch, StaRx 1500, carboxymethylatedstarch), cellulose derivatives, polyvinylpyrollidones, Veegums,polyethylene glycols, sugars, e.g., sucrose and lactose, sodium alginateand waxes.

The fillers used to bulk up a drug tablet or other form also should notinterfere with the tablet's dissolution. Numerous fillers include thestarch derivatives, sugars (e.g., lactose and sucrose), sorbitol,mannitol, cellulose derivatives and their inorganic salts, corn starch,Starch 1500, calcium phosphate, and Avicel.

Likewise, lubricants aid in the machining of a drug tablet. Every tabletneeds a lubricant so that it will be ejected from the machine die withminimum force. However, the lubricant also must not interfere with thedissolution of the tablet. Lubricants include waxes, fatty acids, sodiumsalts of fatty acids and stearates.

The invention is illustrated by the following non-limiting examples.

EXAMPLE 1 Polymerized And Non-Polymerized Lipid Suspensions OfPaclitaxel

6 milligrams of paclitaxel were added to 55 milligrams of polyethyleneglycol 300 and 1.2 milligrams of poloxamer 188 and crystals weredissolved by mixing. At 45° C., 62 milligrams of soybean lecithin andsoybean oil were mixed to which was added 3 milligrams of DODPC, whichdissolved in the lecithin-soy oil. Paclitaxel-PEG was added in toto tothe soy oil, forming a clear oil. In addition, 2 milligrams of HHP(2-hydroxy-4′-(2-hydroxyethoxy)-2-methylpropiophenone, Aldrich) wereadded as a polymerization initiator. The mixture was divided. Oneportion was left as an unpolymerized control. The other portion wasexposed directly to ultraviolet light at 365 nanometers to polymerizethe DODPC in the hydrophobic phase. Polymerization of DODPC wasmonitored by measuring the decrease of absorbance at 254 nanometers. Thehydrophobic phase was polymerized until approximately 30% of themonomers had been reacted. The polymerized hydrophobic phase and theremaining portion not exposed to ultraviolet light were weighed anddispersed into approximately 2× the weight of the hydrophobic phase inwater in which PEG 300 (10% w/v) had been dissolved and Xanthan Gum(0.3% w/v) to form lipid suspensions (formulations D, Table 2). Theunpolymerized lipid spheres were divided into two portions: to oneportion was added HHP and polymerized to 30% completion by exposure toultraviolet light at 365 nanometers. TABLE 1 Composition of PaclitaxelLipid Particles Lipid Particle Formulation (mg)¹ Component D E F GPaclitaxel 6 6 48 25 Perillyl alcohol — 131 144 126 Pluonric F68(poloxamer 188) 1.2 0.65 2.4 1.3 Polyethylene glycol 300 188 185 168 75DODPC 3 3 28 22 Lecithin 62 62 67 76 Soy Oil 62 62 68 73 Xanthan Gum 3 32.4 3.2 Water 653 617 490 631¹Formulations D-G were both polymerized and unpolymerized

EXAMPLE 2 Lipid Particles With Perillyl Alcohol As A Solvent ForPaclitaxel

Formulations E, F, and G (Table 2) were made similarly to formulations Dwith the exception that perillyl alcohol and PEG 300 was used tosolubilize paclitaxel. Polymerization Formulations E-G were prepared asdescribed for Formulation D. More paclitaxel was fully solubilized inthe perillyl alcohol formulations than ones without.

EXAMPLE 3 Stability Of Paclitaxel Lipid Suspensions

Particle size was evaluated for Formulation D, E, F and G. Uniformdistribution of particles with an average radius of approximately 100nanometers was observed upon particle formation. Particle size stabilitywas maintained at 4° C., at room temperature, and at 42°. for allparticle formulations. Paclitaxel crystals were not observedmicroscopically in any of the formulations.

EXAMPLE 4 Bioavailability Of Paclitaxel Following IntraduodenalAdministration Of Lipid Particles In Rats

Sprague-Dawley rats (approximately weighing 220 grams each) werecatheterized surgically with jugular and duodenal catheters. Each groupof rats, 3 animals per group, were given 6 mg/kg of paclitaxel informulation D. Blood samples were collected at 0, 20, 40, 60, 90, 120,and 240 minutes following administration of the formulations. The time 0blood collection was obtaintd approximately 15 minutes beforeexperimental application of formulations. Plasma samples were analyzedby a solid phase extraction of paclitaxel followed by HPLC.Pharmacokinetic parameters were calculated from the data using WinNonLinsoftware (Pharsight). Approximately 100 ng/ml peak plasma concentrationsof paclitaxel was observed in all rats.

EXAMPLE 5 Tumor Regression By Oral Administration Of Paclitaxel As ALipid Suspension

Athymic nude mice are injected subcutaneously with approximately 10⁷MDA-MB-231 cells. Tumors develop at the injection site until they areapproximately 100 mm³ in size. Mice are treated by intraperitonealinjection of cremophor paclitaxel (commercial formulations from BristolMyers Squibb consisiting of ethoxylated Castor oil and ethanol todissolve paclitaxel) or cremophor alone as controls. Subject mice aregiven doses of paclitaxel as lipid suspensions by oral gavage once aday. Tumor size is measured and proportion of mice with tumor regressionis measured.

EXAMPLE 6 Effect Of Lipid Suspensions On Human Breast Cancer Cell Lines

Human breast cancer cell lines are implanted subcutaneously into nudemice. Three human cell lines, MCF-7, BT-20, and MDA-MB-231 are used.Tumors are harvested and cells are grown in RPMI supplemented with fetalbovine serum (10%), ampicillin (100 micrograms per ml), streptomycin,(100 micrograms per ml), and glutamine (0.3%). The cells are grown toapproximately 80% confluence and treated with paclitaxel in Cremophor,Cremophor solution alone, dilution of lipid suspension formulationswithout paclitaxel, or paclitaxel in lipid suspension formulations.Viable cells are determined at times after addition by enumeratingproportion of living cells by dye exclusion technique using tetrazoliumblue.

1. A particle composition that comprises (a) a hydrophobic phase; (b) asurfactant; (c) a biologically active agent; and (d) a polymericstabilizer.
 2. The particle composition of claim 1 wherein thehydrophobic phase comprises an oil or mixture of oils.
 3. The particlecomposition of claim 2 wherein the oil is a triglyceride.
 4. Theparticle composition of claim 1 wherein the hydrophobic phase comprisesa triglyceride that is liquid between 10° C. and 45° C.
 5. The particlecomposition of claim 1 wherein the hydrophobic phase comprises atriglyceride that is liquid between 10° C. and 70° C.
 6. The particlecomposition of claim 1 wherein the surfactant is selected from the groupconsisting of an alkyl glycerolphosphoryl choline, a polyoxyethylenepolymer, a block copolymer of polyoxyethylene and polyoxypropylene, andan ethoxylated glycerol ester.
 7. The particle composition of claim 1,wherein the surfactant comprises one or more glycerol fatty acid estersor hydrophilic derivatives thereof.
 8. The particle composition of claim7, wherein the fatty acid esters have a length of about 6 to about 12carbon atoms.
 9. The particle composition of claim 1, wherein surfactantcomprises a monoglyceride, diglyceride or mixture thereof.
 10. Theparticle composition of claim 1, wherein the surfactant comprises amonoglyceride or a hydrophilic derivative or analog thereof.
 11. Theparticle composition of claim 1, wherein the surfactant comprises adiglyceride or a hydrophilic derivative or analog thereof.
 12. Theparticle composition of claim 1, wherein the surfactant comprises amonoglyceride or diglyceride mixture.
 13. The particle composition ofclaim 6, wherein the surfactant further comprises a diglyceride or ahydrophilic derivative or analog thereof.
 14. The particle compositionof claim 1, wherein the particle suspension further comprises a solventselected from the group consisting of water, glycerol, sorbitol,mannitol, propylene glycol, ethylene glycol, polyethylene glycol ormixtures thereof.
 15. The particle composition of claim 1, wherein theparticle suspension further comprises a solvent selected from the groupconsisting of a monoterpene or a derivative thereof.
 16. The particlecomposition of claim 15, wherein the monoterpene is perillyl alcohol,perillic acid or d-limonene.
 17. The particle composition of claim 10,wherein the monoglyceride is derivatized by a polyoxyethylene polymer.18. The particle composition of claim 10, wherein the monoglyceride isderivatized by a polyoxyethylene polymer from about 200 to about 10,000in molecular weight.
 19. The particle composition of claim 10, whereinthe monoglyceride is derivatized by a polyoxyethylene polymer from about200 to about 4,000 in molecular weight.
 20. The particle composition ofclaim 11, wherein the diglyceride is derivatized by a polyoxyethylenepolymer.
 21. The particle composition of claim 3, wherein at least oneof the fatty acid esters of said triglyceride is derivatized with aceticacid, citric acid, lactic acid, succinic acid, tartaric acid or mixturesthereof.
 22. The particle composition of claim 7, wherein at least oneof the fatty acid esters is a caprylic acid or capric acid.
 23. Theparticle composition of claim 1, wherein the polymeric stabilizer is anatural polymer, a synthetic polymer or a mixture thereof.
 24. Theparticle composition of claims 1 or 23, wherein the polymeric stabilizeris a polymerizable fatty acid or phospholipid.
 25. The particlecomposition of claim 24, wherein the polymeric stabilizer is ODP or OPS.26. The particle composition of claim 24, wherein the polymer is formedfrom a polymerizable fatty acid monomer.
 27. The particle composition ofclaim 1 wherein said composition is in the form of a polymer, saidpolymer formed by interfacial ionic polymerization with water or otherinitiators.
 28. The particle composition of claim 23, wherein saidcomposition is in the form of a polymer, said polymer formed bycondensation of cyanoacrylates, including alkylcyanoacrylates.
 29. Theparticle composition of claim 23, wherein the polymer is formed fromcondensation of ethyl 2-cyanoacrylate.
 30. The particle composition ofclaim 23, wherein the synthetic polymer is selected from the groupconsisting of polylactide, polyglycolide, a mixture of polylactide andpolyglycolide, a polycaprolactone,a polyortho esters, polysebacic acid,polyfumaric acid, polyamides, polycarbonates, polyalkylenes,polyacrylamides, poly(hydroxy acids), polyanhydrides, polyorth oesters,polyacrylate, polyvinyl alcohols, blends or copolymers thereof.
 31. Theparticle composition of claim 1, wherein the particle is suspended in aaqueous phase comprising an additive selected from:the group consistingof a suspending agent, buffering agent, a tonicity agent, an oxidizingagent, a reducing agent, an antimicrobial agent, a preservative, astabilizing agent, or a mixture thereof.
 32. The particle composition ofclaim 1, wherein the hydrophobic phase is present in an amount fromabout 0% to about 50% by weight of the particle composition.
 33. Theparticle composition of claim 1, wherein the hydrophobic phase ispresent in an amount from about 0% to about 50% by weight of theparticle composition.
 34. The particle composition of claim 1,wherein.the hydrophilic phase is present in an amount from about 10% to25% by weight of the particle composition.
 35. The particle compositionof claim 1, wherein the surfactant has an HLB value of about 1 to about45.
 36. The particle composition of claim 1, wherein the surfactant hasan HLB of about 1 to about
 20. 37. The particle composition of claim 1,wherein the biologically active molecule is a prophylactic ortherapeutic agent.
 38. The particle composition of claim 1, wherein thebiologically active molecule is a diagnostic agent.
 39. The particlecomposition of claim 37, wherein the therapeutic agent is a taxane or ananalog thereof.
 40. The particle composition of claim 1 or 37, whereinthe therapeutic agent is paclitaxel.
 41. The particle composition ofclaim 1, wherein the biologically active molecule is a topisomeraseinhibitor.
 42. The particle composition of claim 1, wherein thebiologically active molecule is more than about 0.1 mg/ml soluble in thehydrophobic phase.
 43. The particle composition of claim 1, wherein thebiologically active molecule is more than about 1 mg/ml soluble in thehydrophobic phase.
 44. The particle composition of claim 1, wherein saidcomposition is in the form of a suspension.
 45. The particle compositionof claim 44, wherein the particle size of said suspension is in therange of about 10 nm to 10,000 about nm.
 46. The particle composition ofclaim 44, wherein the particle size of said suspension is in the rangeof about 10 nm to about 1,000 nm.
 47. A method for preventing, treatingor ameliorating one or more symptoms associated with a disease ordisorder in an animal comprising administering to an animal acomposition according to any of claims 1-23, 25-39, 41-44, or
 45. 48. Amethod for preventing, treating or ameliorating one or more symptomsassociated with a disease or disorder in an animal comprisingadministering to an animal a composition according to claim
 24. 49. Amethod for preventing, treating or ameliorating one or more symptomsassociated with a disease or disorder in an animal comprisingadministering to an animal a composition according to claim
 40. 50. Amethod for diagnosing a disease or disorder in an animal comprisingadministering to said animal a composition according to any one ofclaims 1-23, 25-39, 41-44, or
 45. 51. A method for diagnosing a diseaseor disorder in an animal comprising administering to said animal acomposition according to claim
 24. 52. A method for diagnosing a diseaseor disorder in an animal comprising administering to said animal acomposition according to claim
 40. 53. The method according to any oneof claims 47-50, or 51 wherein the composition is administered as acapsule, soft elastic gelatin capsule, caplet, aerosol, spray, solution,suspension, emulsion, sachet, tablet, capsule, powder or granules. 54.The method according to any one of claims 47-52, or 53, wherein saidmammal is a human.
 55. The method according to any one of claims 47-52,or 53, wherein the composition is administered orally.
 56. The methodaccording to any one of claims 47-52, or 53, wherein the polymericstabilizer is a natural polymer, a synthetic polymer or a mixturethereof.
 57. The particle composition of claim 41 wherein thetopoisomerase inhibitor is selected from the group consisting ofetoposide, camptothecin, topotecan or a derivative thereof.
 58. A methodfor orally administering paclitaxel to a patient comprisingadministering the composition of claim 40 to a patient at a dose ofpaclitaxel in the range of about 10 mg/m² to about 1,000 mg/m².